The present invention relates to a method for producing an analysis element spreading layer, which is implemented for the purpose of absorbing a liquid to be analyzed, e.g., a body fluid, in a preferably optical analysis element and distributing it uniformly over a liquid outlet side of the spreading layer. The invention also relates to a spreading layer, which is a porous membrane. Such a spreading layer can be part of an analysis element, which is primarily used for medical analyses. An important area of application of the invention is the assaying of very small sample volumes, which are typically obtained through a puncture in the skin of the patient. The body fluid sample quantities are preferably less than 1 μl.
The analysis is performed using a reagent system, which preferably comprises multiple reagents and auxiliary materials, which are integrated in the analysis element. The reaction of the reagents with an analyte contained in the body fluid results in a measured variable, which is characteristic for the desired analytical result and can be measured on the analysis element. The invention is especially oriented to so-called optical analysis systems and analysis elements, in which the characteristic change of the analysis element, which is a measure of the quantity and/or the presence of an analyte, is optically measurable. The reaction of the body fluid in the analysis element typically results in a change of the color in an analysis layer, which is part of the analysis element and is also designated as a detection layer. The color change of the detection layer is photometrically measured in the wavelength range from approximately 300 nm to approximately 1400 nm. In addition to colorimetric systems, other systems which can be optically evaluated are also known, for example, systems based on fluorescence measurements.
Analysis systems in numerous variants are typical for the qualitative and quantitative determination of various analytes. Systems for determining the glucose concentration in the blood of diabetics have particularly great significance. The invention is particularly suitable for such systems, but it is not restricted thereto. Such systems are performed by the patient himself, for example, for monitoring the health status of the patient (“home monitoring”). Therefore, they must be simple to operate, as small as possible, and robust. Since multiple measurements and therefore also multiple punctures in the skin must be performed daily for a reliable observation of the health status, the quantity of blood removed is to be as small as possible, so that it can also be performed with low puncture depths and therefore with a low-pain puncture.
Optical analysis elements typically have a support structure, which comprises plastic and is often an oblong plastic strip (“test strip”). A so-called test layer, which comprises at least a part of the reagents and the detection layer, adjoins the support layer of the analysis element. The test layer can comprise one or more layers, which are in fluid contact with one another and typically run parallel to one another and to the support layer. Reagent-containing layers (“reagent layers”) of the test layer often comprise an absorbent porous layer material, such as paper or nonwoven material, in which the reagents are impregnated.
In addition to the absorbent porous materials, there are also test layers in which at least one reagent layer is applied to a suitable transparent carrier material by coating. To produce such a coated test layer (CTL), which is designated as a reagent film, reagents mixed into a thickener are applied in the form of a more or less viscous coating compound as a thin film to the carrier material and dried. Upon contact of the reagent film with the aqueous sample liquid, the reactions with the reagents which are required for the analysis occur. For this purpose, the reagent film is typically absorbent and/or swellable and/or soluble. Such systems require a liquid retention structure, which is in fluid contact with the reagent film and uniformly distributes a liquid.
The precision of the analytical results in optical analysis systems and optical analysis elements is substantially dependent on the homogeneity of the coloration in the detection layer of the test layer. In order to reduce inhomogeneities, it is typical to provide a spreading layer adjacent to the test layer of the analysis element, since a uniform distribution of the sample liquid to be analyzed over the test layer is a requirement for a reliable and informative analysis. A liquid outlet side of the spreading layer, which faces toward the test layer, is in fluid contact with a liquid inlet side of the test layer. The spreading layer is implemented so that a liquid penetrating therein is uniformly distributed on its liquid outlet side. During the formation of the spreading layer, materials are used in which a rapid propagation of the body fluid sample occurs over the entire area of the layer. The propagation typically occurs through capillary activity of the spreading layer. Therefore, loose fiber composite structures are suitable, in particular fabrics, papers, or nonwoven materials made of hydrophilic or hydrophilized threads and fibers. Examples of such analysis elements are found in DE 2118455, Boehringer Mannheim GmbH, published Sep. 21, 1972 (see also, U.S. Pat. No. 3,802,842, Lange et al., issued Apr. 9, 1974) and DE 19629657, Boehringer Mannheim GmbH, published Jan. 29, 1998 (see also U.S. Pat. No. 5,846,837, Thym et al., issued Dec. 8, 1998).
In order to ensure reproducible color changes in the detection layer of the analysis element in optical analysis systems, a certain minimum height of a liquid layer having the analyte is necessary, which adjoins the detection layer of the analysis element. The liquid layer thickness is frequently between 50 μm and 200 μm, in particular in the case of enzymatic tests. It is ensured by the accordingly dimensioned spreading layers.
In the case of the processing of small sample volumes less than 1 μl, the known spreading structures have proven to be disadvantageous, however. Structures made of a fine fabric, which are knitted or woven by threads or fibers, have the disadvantage for very small test layers and test layer zones, whose lateral extension of the measuring spot in the test layer plane is in the range of 100 μm to 300 μm, that the mesh width is also in the range of 100 μm. This causes an inhomogeneous formation of the coloration based on the reaction in the detection layer, since the diffusion of analyte from the supernatant sample into the chemical layer is obstructed below a thread. If the threads are processed into a mesh fabric having smaller mesh widths, an excessively small free volume, in which liquid can be absorbed, results in comparison to the total volume of the spreading structure. In addition, the thickness of the thread is excessively large in comparison to the mesh width, so that an optical evaluation is no longer possible. The use of thinner threads is precluded because an adequate layer height in the spreading layer cannot thus be ensured.
Papers or nonwoven materials are also not suitable for the processing of small sample volumes, since they are fundamentally inhomogeneous and have a certain “cloudiness” in the case of optical measurement. The inhomogeneities thus caused in the coloration are negligible in large test layer zones and large optical evaluation areas through averaging. However, in small test layers having an extension of the measuring spot of less than 300 μm in the test layer plane, reliable optical evaluations may not be performed.